Organosilanes have been heretofore used to couple various organic moieties to the surface of inorganic materials. Initially, this coupling was achieved with simple organosilane monomers so that the organosilane was reacted with an inorganic material with the monomer being attached to the inorganic surface through varying numbers of Si--O--Si bonds. The organic portion of organosilane monomers coupled in this manner imparts organic character to the inorganic material and is the basis for the preparation of bonded phase liquid chromatography supports.
Such supports, however, were not completely acceptable since the bonding reaction proved to be slightly reversible. During the elution of columns with thousands of volumes of mobile-phase, soluble organosilane monomers were eluted from the column, and, as the stationary phase eroded from the surface, the chromatographic properties of the column changed.
In addition, it was observed that when the organosilane .gamma.-aminopropyl/triethoxysilane was utilized, a relatively poor surface coverage was obtained during the organosilane bonding reaction and a relatively active surface that denatured proteins was left. This problem in attaching primary amine organosilanes was thought to arise from the formation of ion pairs between amines and surface silanoles. When surface silanoles were sequestered in this manner, they were not available for silaxane bond formation. Later when the support was used in chromatographic separations, these ion pairs dissociated and generated an active surface. The problems of organosilanes leakage from the surface and the presence of residual silanoles made it desirable to pursue alternative bonding chemistry.
A coupling technique refinement was later developed consisting of the attachment of carbohydrate polymers to the surface of porous inorganic supports through functional groups on the organosilanes (see, for example, U.S. Pat. Nos. 3,982,299 and 4,029,583). Preformed polysaccharide polymers were attached to the inorganic supports at many sites during the bonding reaction and subsequently stationary-phase groups attached to these immobilized organic polymers.
Due to multiple site attachment, the polymeric support materials overcame the problem of bonded-phase leakage from the surface, but some problems still remain, at least in some instances and/or usages. For example, where the carbohydrate polymer bonded phases have .gamma.-aminopropyltriethoxysilane as the organosilane anchoring group, these supports retained some of the non-specific adsorption noted above.
An alternative approach for attaching polymers to inorganic supports was also developed and is described, for example, in U.S. Pat. Nos. 4,029,583 and 3,808,125, wherein the organic support has first coupled thereto an organosilane monomer containing a functional group capable of being incorporated into a polymeric matrix during a polymerization reaction. When these supports were then coated with a second monomer and the coated layer polymerized into a three dimensional matrix, the functional group of the organosilane was incorporated into the matrix and the polymer layer bonded to the surface at many sites. Stationary phase groups for chromatography were then attached to this polymer matrix either during the course of polymerization or after polymerization had been completed.
While these polymerization reactions for preparing supports proved to be capable of overcoming the problem of inadequate binding of surface silanoles because neutral silanes were used in the initial coating reaction, the coating reactions proved to be difficult to reproduce in the preparation of five and ten micron particle diameter supports.
Another procedure for the preparation of a bonded-phase chromatography support has also been developed. In this procedure, preformed organic polymers were absorbed to the surface of the inorganic support and the adsorbed layer was then cross-linked to form a pellicle, or skin, on the surface (see, for example, U.S. Pat. No. 4,245,005). In this procedure, there is no covalent bonding between the organic polymer and the inorganic surface, the cross-linked polymer layer is simply held in place by adsorption. This procedure, while being highly reproducible and completely blocking surface silanoles, is usable only for preparing anion exchange supports.
Thus, none of the supports and/or processes for forming such supports have proved to be completely acceptable and have, for example, either lacked stability, quality in the support obtained, or versatility in the synthetic forming procedure.